Abstract The increasing world-wide prevalence of obesity is a worrying public health problem and crisis. Coexistence of obesity, hypertension and diabetes significantly increases the risk of kidney dysfunction. In obesity, concentrations of angiotensin II (Ang II) increase, while those of Ang-(1-7) decrease along with activity of angiotensin converting enzyme 2 (ACE2). Neprilysin (NEP) and ACE2 are metallopeptidases that process Ang I and Ang II, respectively into the Mas receptor agonist, Ang-(1-7). The activity of these protective renin angiotensin system (RAS) components counterbalances the detrimental biological effects of the classical Ang II/angiotensin II type 1 receptor (AT1R) RAS axis. Recently published and preliminary data from our laboratory suggests that treatment with AT2R agonist compound 21 (C21) decreases Ang II and increases Ang-(1-7) in the kidney of obese Zucker rats, by increasing expression and activity of ACE2. Furthermore, our in vitro data suggests that AT2R agonist treatment decreases renin activity as well. Entresto?, a dual AT1R blocker (valsartan) and neprilysin inhibitor (sacubitril) recently released to the market was found to be superior to enalapril therapy alone in the PARADIGM HF trial, resulting in its indication for heart failure with reduced ejection fraction. The benefits of Entresto? are accredited to its ability to increase concentrations of atrial natriuretic peptide (ANP), another substrate of NEP, while blocking the increasing concentrations of Ang II from acting on AT1R. Yet, Entresto? was found to cause an increase in the urinary albumin/creatinine ratio, an indicator of kidney injury. This knowledge along with our preliminary results led to our hypothesis that Ang-(1-7) level in obese kidney is decreased by NEP inhibition, with concurrent increase in Ang II. Combination therapy with C21 increases Ang- (1-7) and attenuates Ang II, by increasing ACE2 activity and expression. Moreover, C21 with NEP inhibition produces an additive effect on increasing ANP levels in obese kidney protecting function and structural integrity of the kidney. To test this hypothesis, Aim 1 is directed to determine the basal levels and basic metabolism of Ang peptides and ANP in obese kidney. Aim 2 will determine that combined treatment (short-term) with NEP inhibitor and AT2R agonist enhances ACE2 activity and Ang-(1-7) levels, and attenuates Ang II levels and ANP degradation in obesity. Aim 3 will determine that chronic combination therapy with a NEP inhibitor and AT2R agonist is reno-protective in salt-induced hypertension in obesity and is superior to Entresto? (ARB + NEP inhibitor). Female obese Zucker rats are included in aim 3 to investigate the sex specific outcomes of our novel combinatory approach and to determine if it is equally effective in both male and female obese Zucker rats. To accomplish these aims, Ang peptides will be quantified utilizing our new liquid chromatography tandem mass spectrometry method; biochemical, histological and hemodynamic approaches to study renal function/injury and telemetry for monitoring blood pressure. The proposed work will significantly contribute to understanding Ang metabolism in obesity and impact future clinical perspectives for a novel reno-protective therapeutic approach.